The present invention relates to analytical chemistry and, more particularly, to centrifuge-based automated sample treatment systems. A major objective of the present invention is to provide rapid and fine temperature control during a series of sample treatments in a centrifuge-induced supergravity field.
The standard of living in modern societies has been greatly enhanced by advances in chemical, biological, and medical sciences. These fields all involve the separation of samples into constituent components that may then be processed to aid in their identification and/or quantification. The centrifuge is an important instance of instrumentation used to separate sample components.
A simple centrifuge has a centrifuge rotor that is spun, e.g., by a motor. Typically, a liquid chemical sample spins with the rotor. The spinning liquid sample components are subjected to a centrifugal force (F=mxcfx892r) proportional to their mass, their distance from the centrifuge spin axis, and the square of the spin rate. The effect of the centrifugal force is much like the effect of gravity-liquid components are separated according to their relative densities. However, unlike gravity, the centrifugal force is readily controlled, e.g., by controlling the spin rate. Thus, a centrifuge can generate centrifugal forces orders of magnitude greater than gravity at the earth""s surface. Generally, the xe2x80x9csupergravityxe2x80x9d conditions of a centrifuge are much more effective than gravity in separating sample components.
In addition, the supergravity conditions afforded by a centrifuge can be used to overcome liquid surface effects that might otherwise impede sample movement. Accordingly, centrifuges that can control the tilt of a chemical-processing unit relative to the centrifugal force can be used for pouring, mixing, filtering, and facilitating chemical reactions. Furthermore, tilting can be used to control liquid movement among multiple processing stations of a chemical-processing circuit so that a series of processes can be implemented without manual intervention. Thus, a centrifuge with tilt control can automate sample processing conventionally performed manually by chemists.
Independent control of centrifuge spin rate and tilt action is disclosed in U.S. Pat. No. 4,814,282 to Holen et al. Tilt of a chemical-processing circuit is used to transfer liquid from one station to another under the influence of centrifugal force. A tilt-drive assembly, including motor and drive chain, is attached to the centrifuge rotor so that it rotates therewith. Power is delivered to the tilt-drive motor via slip rings, which tend to wear out as they are not generally designed to operate at centrifuge speeds. In this approach, any sensors used to track tilt would also rotate at high speeds, further complicating operation. In addition, centrifuge forces are applied to the tilt motor and the drive train. For example, a 1-pound motor must withstand 1000-pound forces in a readily achievable 1000 G supergravity field. Thus, there are a number of robustness issues that can only be addressed with additional complexity and expense.
These robustness issues are mitigated in the centrifuge disclosed in U.S. Pat. No. 5,089,417 to Wogoman. In the Wogoman centrifuge, a holder for a chemical-processing circuit snaps from a first tilt orientation to a second tilt orientation when the centrifuge exceeds a predetermined rotation rate. Similarly, the first tilt orientation is resumed when the centrifuge spin rate falls below the threshold rate. Thus by increasing and decreasing the centrifuge spin rate, sample movement between reaction stations of the chemical-processing circuit can be controlled. However, this approach provides little flexibility in selecting the centrifuge spin rate or tilt angles relative to the centrifugal force. It would be preferable to control the centrifuge rotation and the tilt actions independently.
U.S. Pat. No. 4,776,832 to Martin et al. avoids the need for physical connections to drive a tilt rotor by using inductive motors. The inductive motors include induction rotors that are physically coupled to holders, e.g., for reaction cells, and stationary stators, which are located beneath the centrifuge rotor (wheel). The stators induce eddy currents in the induction rotors, causing them to rotate. No physical connection is required between the stators and the induction rotors, eliminating the need to deliver power through slip rings. On the other hand, the non-physical coupling of drive and induction rotor does not ensure precise and flexible control of sample-container orientation relative to the supergravity field.
Parent U.S. patent application Ser. No. 09/576,690 discloses a coaxial-drive centrifuge in which part of the drive assembly for the tilt motion is coaxial with the centrifuge axis. This arrangement overcomes the robustness limitations of Holen et al., the flexibility limitations of Wogoman, and the precision limitations of Martin et al. A tilt-drive motor provides complete control over tilt without restricting centrifuge rotation rates. The tilt-drive motor is stationary, so electrical coupling is not required to a rotating element. The coupling between the tilt-drive motor and the chemical-processing circuit is mechanical, so there is no problem of precision in tilt control.
The coaxial-drive centrifuge holds the promise for rapid and fully automated sample processing through a series of treatment steps. For example, a polymerase chain reaction (PCR) technique requires many iterations of a series of steps. PCR is used to copy small fragments of doxyribonucleic acid (DNA); the procedure can be iterated so that the amount of DNA grows exponentially. Thus, a limitless amount of DNA sample can be xe2x80x9camplifiedxe2x80x9d from a single DNA fragment. This can allow, for example, multiple parallel destructive analyses to be performed. PCR techniques have accelerated the study of gene functions and gene mappings (e.g., in the Human Genome Project). Generally, PCR is useful in biology, clinical medice, and forensic science.
One variant of PCR , begins with heating a DNA solution (e.g., to 90xc2x0 C.) so that individual strands separate. Then the DNA solution is cooled (e.g., to 50-60xc2x0 C.), allowing oligonucleotide primers to bind to the separated DNA. Then the temperature is raised (e.g., to 70xc2x0 C.) so that polymerase can copy the DNA rapidly. These three phases, melting, annealing, and extension, can be iterated so that the amount of DNA grows exponentially. Typically, the DNA sample remains in a container that is heated and cooled by using temperature controlled baths. The time for the temperature to transfer through the sample container wall is a limiting factor in the rate at which the PCR reaction can be iterated.
Tilt-capable centrifuges with multi-chamber chemical-processing circuits could be used so that each PCR step can be conducted in a dedicated station. Each station can be kept at the temperature associated with one step, e.g., melting, annealing, and extension. Changes in container orientation between steps can be used to move the sample from station to station to automate the processing.
However, the promise for rapid and fully automated chemical-sample processing faces a challenge in temperature control. Typically, different temperatures are required for different sample treatments. The entire centrifuge can be temperature controlled, but then it is difficult to change temperatures rapidly. At best, slow temperature changes delay processing throughput; at worst, slow temperature changes can be incompatible with certain treatment requirements. Local resistive heaters can provide rapid heating. However, delivering electrical power to a rotating chemical-processing circuit for heat control faces linkage challenges as discussed above with respect to Holen et al.
Moreover, sample temperature should be monitored to provide precise closed-loop control thereof. Once again, electrical connections to rotating elements are preferably avoided. What is needed is a system that provides for rapid and precise temperature control to a chemical-processing circuit in the context of a centrifuge.
The present invention provides a centrifuge with inductive temperature control of a tiltable chemical-processing circuit. The centrifuge has a centrifuge rotor, a centrifuge-drive assembly, a chemical-processing unit, a tilt-drive assembly, and an inductor. The tilt-drive assembly is mechanically coupled to the chemical-processing unit to provide for precise and flexible control of tilt relative to the centrifugal force provided by the centrifuge. The chemical-processing unit includes a receptor in which eddy currents are generated when exposed to the alternating magnetic field generated by the inductor. The eddy-current energy is dissipated as heat due to resistance in the receptor.
Energy transfer between the inductor and the receptor is contactless. Accordingly, the inductor can be xe2x80x9cstationaryxe2x80x9d in the sense that it does not rotate with the centrifuge rotor or the chemical-processing circuit. Thus, electrical power can be supplied to the inductor through standard cabling. So that power is not wasted, and more tightly directed to a small zone on the rotor, the alternating magnetic field can be preferably generated only at times that are at least in part a function of the centrifuge rotor orientation.
The chemical-processing unit can include an optical temperature sensor. This can be read by a stationary optical reader as the chemical-processing circuit rotates by. The optical reader can be powered via standard electrical cabling and its signal output can be provided to a controller that regulates the temperature of the sample. The temperature at which the sample is to be maintained depends on the treatment, which can correspond to the station or container location of the sample. Thus, the maintained temperature can be determined as a function of the tilt orientation of the chemical-processing circuit relative to the centrifugal force applied by the centrifuge.
The basic method provided for by the present invention involves spinning a chemical-processing unit about a centrifuge axis so that a centrifugal force is applied to the chemical sample. Movement of the sample within the chemical-processing unit is effected by titling the chemical-processing unit relative to the centrifugal force. Heating of the sample is achieved by inducing eddy currents in the chemical-processing unit using an inductor. Preferably, the inductor generates the eddy currents only when a receptor is aligned with the inductor, as determined by the centrifuge orientation. Regulation of sample temperature can be achieved by contactlessly reading a thermo-sensor included with the chemical-processing unit.
If a chemical-processing circuit (a multi-station chemical-processing unit with channels permitting sample flow between stations) is used, the method of the invention provides for automated processing sequences. The chemical-processing circuit can be oriented relative to the centrifugal force so that the chemical sample is maintained at the first processing station. Once the first treatment is complete, the chemical-processing circuit can be tilted so that the sample flows to a second station for a second treatment.
Where the different treatments require different temperatures, each station can include its own thermo-sensor. The contactless optical reader can read whichever thermo-sensor corresponds to the present location of the sample. A controller can vary inductor pulse widths to control the rate of heating as appropriate to minimize deviations of the actual sample temperature from the target sample temperature.
In a more specific method of the invention, a sample is inserted into a multi-station chemical-processing circuit. Centrifugal force is applied with the chemical-processing circuit oriented so that the sample is held in the first station. In the meantime, the temperature of the sample is regulated using a sensor in the container so that a first temperature is maintained. When the first treatment is completed, the orientation of the chemical-processing circuit relative to the centrifugal force is changed so that the sample moves to the second station. Then the sample is maintained in the second station while the second treatment is applied at a second temperature maintained through regulation response to the sensor.
In the context of PCR, the present invention permits rapid amplification of DNA fragments. For example, each of three stations of a chemical-processing circuit can be dedicated to one of the three phases, melting, annealing, and extension, of the PCR procedure. The chemical-processing circuit can be tilted to pour the DNA sample from station to station. Each station can be kept at the appropriate temperature for the corresponding phase of the PCR procedure. The chemical-processing circuit can be rocked back and forth to agitate the DNA sample to facilitate uniform temperature changes. Thus, the invention provides for rapid automated PCR reactions. Reactions with similar requirements for temperature changes and agitation are similarly facilitated by the present invention.
The present invention provides for rapid and fully automated multi-treatment sample processing with rapid and precise contactless sample temperature control. The chemical-processing circuit is not encumbered either by cables to supply heat or to power a temperature sensor, or to provide feedback to a controller. These and other features and advantages of the invention are apparent from the description below with reference to the following drawings.